The invention relates to a method for culturing biological cells, in particular a method for forming biological cell aggregates, and to a cell culturing device.
It is generally known to culture biological cells outside an organism (in vitro culturing). Typically the cells form a cell culture on a plastics or glass substrate in a culturing medium. For example, a growth and/or differentiation of the cells is provided on the substrate. In numerous cell biology methods the task is that of removing groups of cells from the cell culture, for example in order to subject these groups to further culturing or to an analysis. In conventional standard cell biology methods, groups of cells are removed from a cell culture using mechanical means (e.g. using a separating tool) or using chemical means (e.g. by enzymatic degradation).
With in vitro culturing on a substrate, the cells typically form a bed of cells having few layers of cells or having only a single layer of cells (monolayer). The formation of thin layers presents a problem if large groups of cells are required for study or further culturing. In this case the cells must be separated from the substrate over a relatively large surface area. This can lead to undesirable biochemical reactions which impair the subsequent method steps.
For specific cell types or culturing media, growth beyond the monolayer can be achieved in individual cases. The cells form three-dimensional cell aggregates. For example, in the adherent state stem cells form cell aggregates (what are known as “organoid bodies”, see C. Kruse et al. in Appl. Phys. A, vol. 79, 2004, p. 1617-1624; and C. Kruse et al. in Ann. Anat., vol. 188(6), 2006, p. 503-517).
The above-mentioned standard cell biology methods can likewise be used to separate three-dimensional cell aggregates from a cell culture. In practice, however, the problem with this is that the cell aggregates are not uniformly formed on the substrate. On the conventionally used substrates, the cell aggregates form an inhomogeneous distribution, in particular in respect of the geometric arrangement and size of the cell aggregates. Furthermore, the typically spontaneously formed cell aggregates are at different stages of development. Thus a cell aggregate can include specific differentiated cells, while a younger cell aggregate does not include these cells. In practice, however, there is often interest in removing from a cell culture, for subsequent method steps, a plurality of cell aggregates having characteristics that are as similar as possible, in particular having, as far as possible, the same size and shape and the same stage of development. Thus hundreds or thousands of cell specimens of the same type are required for high-throughput test methods where e.g. the biological action of a pharmacological substance is studied.
A further limitation of the conventional culturing techniques relates to the shape of the cell aggregates. Cell aggregates are combined into larger aggregate formations for the provision of tissue models for study purposes or for the formation of implants in what is known as “tissue engineering”. There is thus interest in the cell aggregates having a specific shape to make it easier to combine the aggregate formation. Using the conventional mechanical means, however, cell aggregates can be shaped only with considerable effort, while shaping is virtually impossible where cells are enzymatically degraded from a cell culture.
Thus only small groups of cells which are too small for further method steps, or cell aggregates having undesirably heterogeneous characteristics can be obtained from a cell culture using the conventional standard cell biology methods.
It is also known to culture biological cells on porous substrates (e.g. EP 0 759 064 B1, WO 97/00314). A porous substrate allows the cells to be controlled, e.g. with an active substance, from the substrate side. Conventional porous substrates have such small pore sizes that a cell covers over a pore (see e.g. DE 10 2004 062 216 A1). Handling of the cells on porous substrates poses the same problems as those mentioned above in relation to the standard methods of in vitro culturing.
The objective of the invention is that of providing an improved method for culturing biological cells, a method with which the problems and limitations of conventional methods are overcome. The objective of the invention is also that of providing an improved device for culturing biological cells, a device with which disadvantages of conventional culturing techniques are overcome.
These objectives are achieved by means of a method and/or a cell culturing device of the invention.